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Simultaneous Detection of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis by a Rapid PCR Method

Department of Biochemistry and Biotechnology, A.I. Virtanen Institute

J.H. Meurman

Faculty of Dentistry, University of Kuopio, PO Box 1627, FIN-70211 Kuopio, Finland

P. Väisänen

Faculty of Dentistry, University of Kuopio, PO Box 1627, FIN-70211 Kuopio, Finland

P. Alakuijala

Faculty of Dentistry, University of Kuopio, PO Box 1627, FIN-70211 Kuopio, Finland

A. Korhonen

Faculty of Dentistry, University of Kuopio, PO Box 1627, FIN-70211 Kuopio, Finland

H. Torkko

Oral Microbiology Service Laboratory, Institute of Dentistry, University of Helsinki, Helsinki, Finland

J. Janne

Department of Biochemistry and Biotechnology, A.I. Virtanen Institute

The identification of periodontal pathogens by conventional methods is time-consuming and difficult. Therefore, a multiplex PCR method for simultaneous detection of Actinobacillus actinomycetemcomitans (A.a.) and Porphyromonas gingivalis (P.g.) was developed for rapid and easy determination of these risk-indicator bacteria in human periodontal disease. The PCR primers were designed to hybridize to various regions of 16S rRNA genes, and a hot-start technique was used to obtain maximum sensitivity and specificity. This method can detect both of these bacteria in subgingival plaque samples at concentrations as low as 5 to 50 cells per sample. The sensitivity, however, was even 10 times better when the bacteria were analyzed in a water suspension. Since the only step between sample collection and the actual analysis is a brief centrifugation of the patient sample, the detection can be readily carried out in four hours. The performance of the method was studied with 36 patient samples. The results showed that the PCR method detected A.a. (44% vs. 25%, respectively) and P.g. (56% vs. 42%, respectively) more often than the conventional culture in plaque samples. Thus, our multiplex PCR method is rapid and more effective than conventional protocols in detecting these periodontal pathogens.

Key Words: Actinobacillus actinomycetemcomitans • Porphyromonas gingivalis • polymerase chain reaction.

Journal of Dental Research, Vol. 74, No. 11, 1796-1801 (1995)
DOI: 10.1177/00220345950740111301


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